Studying the effects of sidechain mutation on protein stability, pKa shifts, or other properties of a protein using molecular simulations may require the building of an appropriately mutated structure since a crystal structure of the desired mutant may not be available. One could manually replace a residue with another directly by editing the PDB file, but this may lead to an energetically unfavorable position. To obtain a more realstic mutated structure a better approach is to replace the sidechain, then select from possible rotamers found in other protein crystal structures and use some sort of scoring function to select the best (lowest energy) possible rotamer. Fortunately Molden (G.Schaftenaar and J.H. Noordik, "Molden: a pre- and post-processing program
for molecular and electronic structures", J. Comput.-Aided Mol. Design,
14 (2000) 123-134 ) provides a facility to perform this rotamer search that is fast, simple to use, and provides reasonable computationally mutated proteins structures. This page will show the steps to perform the task of preparing a mutated protein structure using molden in step-by-step manner for your convenience.
First, we need a protein crystal structure. I will use bovine alpha-lactalbumin (ALB) for no particular reason (PDBID : 1F6R.pdb).
The crystal structure contains several copies of ALB in one unit, so you must select one of these structures.
You may use the linux commands as follows to select the first model structure (pay attention to the single and double quotes!).
awk '$1=="ATOM"&&$5=="A"' 1F6R.pdb > 1f6r_a.pdb
If you don't know much about awk this command says "look in file 1F6R.pdb, if the first column has the string atom AND the fifth column has the string A copy this to the new file 1f6r_a.pdb
now you can view ALB using molden
molden 1f6r.pdb
To make a mutation click on the button "ZMAT editor"
Now you can see a list of the residues, and a list of relative position of the atoms for each residues
To mutate a residue click on the button for a particular residue, or move the slider until you find the residue you want to mutate.
When you click on a residue a window will pop-up will several options. To mutate, click on "replace"
When you click on "Replace" a window will pop-up with options of what residue you can replace it with. I chose Arginine.
Now ARG has replaced the previous residue, the it is in an unfavorable position. We need to select the best from a rotamer library.
Click on the button corresponding to the mutated residue and the same window as before pops-up.
Click on "best rotamer" and molden search for a better sidechain position
Now we just save. Click on button "Write" and select 'PDB'
You can chose with or without hydrogens.
Then enter the file name to save your newly mutated protein.
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Easier than real chemistry I would guess!